Project summary Mycobacterium tuberculosis (M.tb) causes more deaths worldwide than any other infectious agent and is increasingly characterized by antimicrobial resistance. An effective vaccination strategy to prevent establishment of M.tb infection and progression to tuberculosis (TB) disease is a priority to achieve rapid control of the global TB epidemic, but development and clinical testing of new vaccines is hampered by the lack of immune correlates of protection. Revaccination of adolescents with the only licenced vaccine against TB, Bacille Calmette-Guerin (BCG), partially protected against established M.tb infection, providing the first efficacy signal for a novel TB vaccination strategy in a high-risk population. There is a one-time opportunity to leverage this existing cohort, as well as newly enrolled participants in a larger confirmatory phase 2b clinical trial, to identify and test the durability of immune correlates of protection against established M.tb infection in the lung, which could harbour critically different immune responses compared to peripheral blood. Blood and bronchoalveaolar lavage specimens will be collected to identify and determine longevity of vaccine- induced systemic and pulmonary immune responses that correlate with protection against sustained M.tb infection. The durability of vaccine-induced protection against infection ~6 years after vaccination will also be explored. Novel and robust technologies, such as polychromatic flow cytometry, mass cytometry and systems serology will be applied to study adaptive and innate immune responses in participants enrolled in two BCG efficacy trials at 6 months and ~6 years after vaccination as well as 1 month and 4-5 years after primary M.tb infection. The overarching hypothesis is that BCG vaccination induced multiple arms of the immune system and that a combination of Th1/Th17 and IgA responses is associated with lower risk of established M.tb infection. This approach will: 1) Identify and assess durability of BCG-mediated systemic and pulmonary immune responses in M.tb-uninfected individuals; 2) Define systemic and pulmonary immune correlates of post- M.tb exposure protection against established M.tb infection in participants experiencing transient (protected) or sustained (unprotected) M.tb infection. Identification of vaccine-induced immune responses important for protection against established M.tb infection would enhance our understanding of early containment of M.tb and could inform immune correlates of protection against TB disease. By extending the follow-up of the first BCG revaccination trial and comparing rates of M.tb infection in participants from the BCG and placebo arms, we will also 3) Measure the durability of vaccine-mediated prevention of sustained M.tb infection for at least 6 years post-vaccination, which, if significant, would provide impetus to conduct larger trials testing for efficacy to prevent TB disease in endemic populations.